Borrelia burgdorferi is an obligate parasite, and is maintained in nature through a complex cycle involving both a tick and mammalian host. The transition between these two very different host types requires the ability to rapidly adapt through changes in gene expression. Artificial infection studies have provided information on a number of these genes that may be differentially expressed under conditions similar to that of the vertebrate host. Despite this advance, there remains a fundamental gap in our understanding of the role and function of a number of genes for mammalian host adaptation. Our long-term goal is to identify and characterize the proteins necessary for adaptation of B. burgdorferi to both the tick and mammalian host environments. The overall objective of this application is to identify B. burgdorferi candidate genes required for adapting specifically to the mammalian host using a mouse model. The central hypothesis is that a large number of B. burgdorferi genes of unknown function are adaptively expressed. Furthermore, we hypothesize that a number of genes important for host adaptation are either transiently expressed, or are turned on only during late-stage infection of mice. The rationale for the proposed research is that, once the genes involved in adapting to the host environment have been identified, it will be possible to determine their respective roles in establishing mammalian infection. Thus, the proposed research is relevant to that part of NIH's mission that pertains to developing fundamental knowledge that will potentially help to reduce the burdens of human illness and disability. Guided by the published B. burgdorferi genome sequence and cited work by other groups, our hypothesis will be tested by pursuing two specific aims: 1) Identify genes involved in the adaptation of B. burgdorferi to the mammalian host environment;and 2) Identify late-stage host adaptation factors of B. burgdorferi important for mammalian infection. Under the first aim, we will make use of In Vivo Expression Technology (IVET). This system will utilize a promoter-based genomic DNA library of B. burgdorferi that is designed to allow for the selection of only those promoters that are active exclusively within the mouse host. The second aim will utilize a modified version of the IVET system, termed Recombinase-based IVET (RIVET). This system will also make use of a promoter-based genomic DNA library of B. burgdorferi, but is designed to allow for the selection of only those promoters that are active transiently or during late-stage infection in the mouse host. The proposed work is innovative because it capitalizes on the essential activities of specific B. burgdorferi proteins as a new means of identifying important virulence factors. When applied, the results from the proposed studies are expected to allow the targeting of this system in order to significantly reduce the ability of this pathogen to establish infection in the mammalian host. PUBLIC HEALTH RELEVANCE: The proposed studies are of an important area of Lyme disease research that has applicability to understanding infectivity and pathogenesis by Borrelia burgdorferi. The proposed research has relevance to public health because the resulting discoveries have the potential to fundamentally advance the field of B. burgdorferi adaptive gene expression, and may have broad implications for host adaptation systems in other animal and human pathogens. Thus, the findings are ultimately expected to be applicable to the health of human beings.